Hierarchical self-tolerance to T cell determinants within the ubiquitous nuclear self-antigen La (SS-B) permits induction of systemic autoimmunity in normal mice.
Systemic autoimmune diseases are frequently associated with clustering of high titer autoantibody responses towards nuclear self-antigens. Little is known, however, about the extent of immune tolerance to the target nuclear antigens or the events leading to the complex autoantibody responses that are characteristic of systemic autoimmunity. To address these issues, we have examined the mouse immune response to La autoantigen (mLa) and the homologous human La antigen (hLa), which are components of the La(SS-B)/Ro(SS-A) ribonucleoprotein (RNP) complex targeted in systemic lupus erythematosus and primary Sjögren's syndrome. The findings reveal the presence of hierarchical T cell tolerance involving multiple autodeterminants within the La autoantigen expressed by normal H-2k and H-2a mice. At one end of this spectrum, there was no detectable T or B cell autoimmunity observed in mice that were immunized with the immunodominant mLa287-301 determinant, which differed by a single residue in its core sequence from the homologous but highly immunogenic human La288-302 determinant. Interestingly, the mLa287-301 peptide acted as an altered peptide ligand that specifically antagonized the activation of an hLa288-302-specific T cell hybridoma. In contrast to the tolerogenic mLa287-301 determinant, a range of autoimmune potential was identified among poorly tolerizing, subdominant self-peptides present within mouse La autoantigen. Notably, immunization of normal mice with the autologous subdominant La25-44 and La106-129 determinants resulted in limited or no detectable autoantibody response. In contrast, immunization with the subdominant mouse La13-30 determinant induced a proliferative T cell response associated with the appearance of specific autoantibodies recognizing multiple intrastructural (La) and intermolecular components (Ro) of the murine La/Ro RNP. The findings suggest how diversified autoimmunity might follow initiation of immunity to simple peptide mimics of poorly tolerogenic determinants that are present within ubiquitous self-antigens.


Assessing Swallowing of the Breastfeeding NICU Infant Using Fiberoptic Endoscopic Evaluation of Swallowing: A Feasibility Study.
Premature infants may demonstrate feeding difficulties requiring an instrumental swallowing assessment. Fiberoptic endoscopic evaluation of swallowing (FEES) is one assessment that can evaluate bottle feeding and breastfeeding. This pilot study investigated the safety and feasibility of FEES for neonatal intensive care unit (NICU) infants during breastfeeding. The setting for this prospective, descriptive study was an urban level III NICU that provided care for premature and critically ill infants. Participants were 5 infants recruited from a convenience sample who were at least 37 weeks postmenstrual age, demonstrated feeding difficulties during a bedside feeding and swallowing examination, and were breastfeeding. Each participant received a FEES assessment while breastfeeding (FEES-B). Adverse events and vital signs including prefeeding and postfeeding respiratory rate, heart rate, and oxygen saturation level were recorded to assess safety. Visualization of milk, laryngeal penetration, and tracheal aspiration assessed feasibility. Participants had a mean postmenstrual age of 39.8 weeks. No adverse events, including epistaxis or laryngospasm, were reported; there were no instances of autonomic instability; and there were no statistically significant differences between prefeeding and postfeeding respiratory rate, heart rate, or oxygen saturation level. Two infants successfully breastfed during FEES-B. Human milk was observable, and laryngeal penetration was noted with 1 infant. FEES-B was found to be safe, with limited data supporting feasibility. Infants demonstrated no physiologic instability during FEES-B, and milk was visualized in the pharynx during breastfeeding with 2 infants. Further studies are warranted to assess effectiveness of FEES-B in infants.


Spectroscopic investigations on thin adhesive layers in multi-material laminates.
Three different spectroscopic approaches, Raman linescans, Raman imaging, and attenuated total reflection Fourier transform infrared spectroscopy (ATR FT-IR) imaging were evaluated for the visualization of the thin adhesive layers (3-6 μm) present in polymeric photovoltaic backsheets. The cross-sections of the multilayer laminates in the original, weathered, and artificially aged samples were investigated spectroscopically in order to describe the impact of the environmental factors on the evenness and thickness of the adhesive layers. All three methods were found to be suitable tools to detect and visualize these thin layers within the original and aged polymeric laminates. However, as the adhesive layer is not very uniform in thickness and partly disintegrates upon weathering and/or artificial aging, Raman linescans yield only qualitative information and do not allow for an estimation of the layer thickness. Upon increasing the measuring area by moving from one-dimensional linescans to two-dimensional Raman images, a much better result could be achieved. Even though a longer measuring time has to be taken into account, the information on the uniformity and evenness of the adhesive layer obtainable using the imaging technique is much more comprehensive. Although Raman spectroscopy is known to have the superior lateral resolution as compared with ATR FT-IR spectroscopy, the adhesive layers of the samples used within this study (layer thickness 3-6 μm) could also be detected and visualized by applying the ATR FT-IR spectroscopic imaging method. However, the analysis of the images was quite a demanding task, as the thickness of the adhesive layer was in the region of the resolution limit of this method. The information obtained for the impact of artificial aging and weathering on the adhesive layer obtained using Raman imaging and ATR FT-IR imaging was in good accordance.


A road map to comprehensive androgen receptor axis targeting for castration-resistant prostate cancer.
Gonadal androgen suppression (castration via orchiectomy or gonadotropin-releasing hormone analogues) suppresses circulating testosterone levels but does not achieve adequate androgen ablation within the prostate cancer microenvironment because it does not address adrenal and intratumoral steroid contributions. These residual extragonadal sources of androgens allow prostate cancer cells to survive, adapt, and evolve into castration-resistant prostate cancer (CRPC). The persistent significance of the androgen receptor (AR) axis in CRPC was recently validated by the clinical efficacy of androgen synthesis inhibitors (abiraterone) and novel, second-generation AR antagonists (enzalutamide). The appreciation that conventional therapeutic approaches achieve a suboptimal ablation of intratumoral androgens and AR axis signaling output opens transformative therapeutic opportunities. A treatment paradigm of comprehensive AR axis targeting at multiple levels (androgen synthesis, metabolism, and action) and at all relevant sites (gonadal, adrenal, intratumoral) simultaneously at the time of initiation of endocrine therapy (instead of the current approach of sequentially adding one agent at a time and only after disease progression) deserves examination in clinical trials to explore whether maximal first-line AR axis suppression via combination therapy can achieve maximal induction of cancer cell apoptosis (before they have the chance to adapt and evolve into CRPC) and thus, improve patient outcomes. Cancer Res; 73(15); 4599-605. ©2013 AACR.


Circulating CA125 in patients with peritoneal mesothelioma treated with cytoreductive surgery and intraperitoneal hyperthermic perfusion.
Recent phase I/II trials report encouraging results in selected patients with peritoneal mesothelioma (PM) treated with cytoreductive surgery (CRS) and intraperitoneal hyperthermic perfusion (IPHP). Circulating tumor markers have never been extensively investigated in the management of PM. We assessed the clinical role of markers in a large series of patients with PM undergoing CRS and IPHP. Clinical data on 60 patients with PM operated with the intention to perform adequate CRS (residual tumor nodules <or= 2.5mm) and IPHP were prospectively collected. Marker levels were determined pre-operatively, post-operatively, and routinely during long-term follow-up. Baseline diagnostic sensitivity, accuracy in monitoring response to treatment or tumor progression and prognostic significance were determined. Baseline diagnostic sensitivity was 53.3% for CA125, 0 for CEA, 3.8% for CA19.9 and 48.5% for CA15.3. Forty-six patients underwent adequate cytoreduction and IPHP; gross residual tumor was left after the operation in fourteen. Postoperatively, CA125 became negative in 21/22 patients with elevated baseline levels undergoing adequate CRS and IPHP, while remained elevated in 9/9 patients with persistent macroscopic disease. CA125 became positive in 12/12 patients with elevated baseline levels developing disease progression after adequate CRS and IPHP. Baseline CA125 showed borderline prognostic significance only among patients not previously treated with systemic chemotherapy. CA125 was elevated in the majority of patients with PM in the present series. Serial maker measurements paralleled tumor growth or regression after CRS and IPHP, suggesting the need of further studies to assess the role of CA125 in this clinical setting.


Genetic analysis identifies a function for the queC (ybaX) gene product at an initial step in the queuosine biosynthetic pathway in Escherichia coli.
Queuosine (Q), one of the most complex modifications occurring at the wobble position of tRNAs with GUN anticodons, is implicated in a number of biological activities, including accuracy of decoding, virulence, and cellular differentiation. Despite these important implications, its biosynthetic pathway has remained unresolved. Earlier, we observed that a naturally occurring strain of Escherichia coli B105 lacked Q modification in the tRNAs. In the present study, we developed a genetic screen to map the defect in E. coli B105 to a single gene, queC (renamed from ybaX), predicted to code for a 231-amino-acid-long protein with a pI of 5.6. As analyzed by mobility of tRNA(Tyr) on acid urea gels and two-dimensional thin-layer chromatography of the modified nucleosides, expression of QueC from a plasmid-borne copy confers a Q+ phenotype to E. coli B105. Further, analyses of tRNA(Tyr) from E. coli JE10651 (queA mutant), its derivative generated by deletion of chromosomal queC (queA deltaqueC), and E. coli JE7325, deficient in converting preQ0 to preQ1, have provided the first genetic evidence for the involvement of QueC at a step leading to production of preQ0, the first known intermediate in the generally accepted pathway that utilizes GTP as the starting molecule. In addition, we discuss the possibilities of collaboration of QueC with other cellular proteins in the production of preQ0.


Cl-out is a novel cooperative optogenetic tool for extruding chloride from neurons.
Chloride regulation affects brain function in many ways, for instance, by dictating the GABAergic reversal potential, and thereby influencing neuronal excitability and spike timing. Consistent with this, there is increasing evidence implicating chloride in a range of neurological conditions. Investigations about these conditions, though, are made difficult by the limited range of tools available to manipulate chloride levels. In particular, there has been no way to actively remove chloride from neurons; we now describe an optogenetic strategy, 'Cl-out', to do exactly this. Cl-out achieves its effect by the cooperative action of two different component opsins: the proton pump, Archaerhodopsin and a chloride channel opsin. The removal of chloride happens when both are activated together, using Archaerhodopsin as an optical voltage clamp to provide the driving force for chloride removal through the concurrently opened, chloride channels. We further show that this novel optogenetic strategy can reverse an in vitro epileptogenic phenotype.


Hypoxia promotes acquisition of aggressive phenotypes in human malignant mesothelioma.
Hypoxia is a hallmark of the solid tumor microenvironment and is associated with poor outcomes in cancer patients. The present study was performed to investigate mechanisms underlying the hypoxia-induced phenotypic changes using human malignant mesothelioma (HMM) cells. Hypoxic conditions were achieved by incubating HMM cells in the air chamber. The effect of hypoxia on phenotype changes in HMM cells was investigated by performing in vitro clonogenicity, drug resistance, migration, and invasion assays. Signaling pathways and molecules involved in the more aggressive behaviors of HMM cells under hypoxia were investigated. A two-tailed unpaired Student's t-test or one-way ANOVA with Bonferroni post-test correction was used in this study. Hypoxic conditions upregulated hypoxia-inducible factor 1 alpha (HIF-1α) and HIF-2α in parallel with the upregulation of its target, Glut-1, in HMM cells. In vitro clonogenicity of HMM cells was significantly increased in hypoxic conditions, but the proliferation of cells at a high density in hypoxia was lower than that in normoxic conditions. The expression levels of HIF-2α and Oct4 were increased in hypoxic HMM cells. The percentage of cells with high CD44 expression was significantly higher in HMM cells cultured in hypoxia than those cultured in normoxia. Hypoxia significantly enhanced the resistance of HMM cells to cisplatin, which occurred through cytoprotection against cisplatin-induced apoptosis. While cisplatin treatment decreased the ratio of Bcl-2 to Bax in normoxic condition, hypoxia conversely increased the ratio in HMM cells treated with cisplatin. Hypoxia increased the mobility and invasiveness of HMM cells. Epithelial to mesenchymal transition was promoted, which was indicated by the repression of E-cadherin and the concomitant increase of vimentin in HMM cells. The data illustrated that hypoxic conditions augmented the aggressive phenotypes of HMM cells at the biological and molecular levels. The present study provides valuable background information beginning to understand aggressiveness of HMM in tumor microenvironments, suggesting that a control measure for tumor hypoxia may be an effective therapeutic strategy to reduce the aggressiveness of cancer cells in HMM patients.


Microbial Community Structure in a Serpentine-Hosted Abiotic Gas Seepage at the Chimaera Ophiolite, Turkey.
The surface waters at the ultramafic ophiolitic outcrop in Chimaera, Turkey, are characterized by high pH values and high metal levels due to the percolation of fluids through areas of active serpentinization. We describe the influence of the liquid chemistry, mineralogy, and H2 and CH4 levels on the bacterial community structure in a semidry, exposed, ultramafic environment. The bacterial and archaeal community structures were monitored using Illumina sequencing targeting the 16S rRNA gene. At all sampling points, four phyla, Proteobacteria, Actinobacteria, Chloroflexi, and Acidobacteria, accounted for the majority of taxa. Members of the Chloroflexi phylum dominated low-diversity sites, whereas Proteobacteria dominated high-diversity sites. Methane, nitrogen, iron, and hydrogen oxidizers were detected as well as archaea and metal-resistant bacteria.IMPORTANCE Our study is a comprehensive microbial investigation of the Chimaera ophiolite. DNA has been extracted from 16 sites in the area and has been studied from microbial and geochemical points of view. We describe a microbial community structure that is dependent on terrestrial, serpentinization-driven abiotic H2, which is poorly studied due to the rarity of these environments on Earth.


Complete amino acid sequence of kaouthiagin, a novel cobra venom metalloproteinase with two disintegrin-like sequences.
The primary structure of kaouthiagin, a metalloproteinase from the venom of the cobra snake Naja kaouthia which specifically cleaves human von Willebrand factor (VWF), was determined by amino acid sequencing. Kaouthiagin is composed of 401 amino acid residues and one Asn-linked sugar chain. The sequence is highly similar to those of high-molecular mass snake venom metalloproteinases from viperid and crotalid venoms comprised of metalloproteinase, disintegrin-like, and Cys-rich domains. The metalloproteinase domain had a zinc-binding motif (HEXXHXXGXXH), which is highly conserved in the metzincin family. Kaouthiagin had an HDCD sequence in the disintegrin-like domain and uniquely had an RGD sequence in the Cys-rich domain. Metalloproteinase-inactivated kaouthiagin had no effect on VWF-induced platelet aggregation but still had an inhibitory effect on the collagen-induced platelet aggregation with an IC(50) of 0.2 microM, suggesting the presence of disintegrin-like activity in kaouthiagin. To examine the effects of these HDCD and RGD sequences, we prepared synthetic peptides cyclized by an S-S linkage. Both the synthetic cyclized peptides from the disintegrin-like domain and from the Cys-rich domain) had an inhibitory effect on collagen-induced platelet aggregation with IC(50) values of approximately 90 and approximately 4.5 microM, respectively. The linear peptide (RAAKHDCDLPELC) and the cyclized peptide had little effect on collagen-induced platelet aggregation. These results suggest that kaouthiagin not only inhibits VWF-induced platelet aggregation by cleaving VWF but also disturbs the agonist-induced platelet aggregation by both the disintegrin-like domain and the RGD sequence in the Cys-rich domain. Furthermore, our results imply that the corresponding part of the Cys-rich domain in other snake venom metalloproteinases also has a synergistic disturbing effect on platelet aggregation, serving as a second disintegrin-like domain. This is the first report of an elapid venom metalloproteinase with two disintegrin-like sequences.


A randomized controlled trial to compare the complications of 2 peripheral intravenous catheter-stabilization systems.
An open-label, prospective, randomized, noninferiority study was conducted at a large academic, Magnet-designated, Level I trauma center to compare the peripheral intravenous catheter securement-related complication rates of 2 different stabilization systems. The control stabilization system included the StatLock device with a nonwinged catheter, and the investigational stabilization system included a closed catheter system with a specially designed Tegaderm dressing. Data from 302 subjects indicated that the investigational stabilization system was noninferior or similar to the control stabilization system with respect to the overall securement-related complications. The cost of the investigational stabilization system was approximately 75% of the cost for the control stabilization system.


Prevalence of gingival overgrowth among elderly patients under amlodipine therapy at a large Indian teaching hospital.
To determine the prevalence of amlodipine-induced gingival overgrowth (GO) among elderly subjects attending an Indian teaching hospital and find any association with demographic factors, drug variables, oral hygiene status and gingival inflammation. A cross-sectional pilot study included 157 dentate patients aged 60 years or more, taking Amlodipine for at least 3 months. Data were collected from past medical records and oral examination. Clinical assessment of GO was correlated with patient's age, gender, drug dosage (2.5, 5 or 10 mg/day), duration of drug therapy (3-4, 4-6, 6-12, 12-24 and >24 months) and also with subjects' plaque index and gingival index scores. Eight patients (5.09%) had GO. No statistically significant relation was observed between age (p = 0.79), gender (p = 0.56), drug dosage (p = 0.25) and duration of drug intake (p = 0.62) and prevalence of GO. GO prevalence related highly significantly (p < 0.001) with plaque and gingival index scores. Prevalence of amlodipine-associated GO in the sample of elderly Indian patients was noted higher than that previously reported. Plaque and gingival inflammation were highly correlated with this condition, while demographic characteristics and drug dosage did not relate significantly.


Phylogenetic study and identification of Vibrio splendidus-related strains based on gyrB gene sequences.
Different strains related to Vibrio splendidus have been associated with infection of aquatic animals. An epidemiological study of V. splendidus strains associated with Crassostrea gigas mortalities demonstrated genetic diversity within this group and suggested its polyphyletic nature. Recently 4 species, V. lentus, V. chagasii, V. pomeroyi and V. kanaloae, phenotypically related to V. splendidus, have been described, although biochemical methods do not clearly discriminate species within this group. Here, we propose a polyphasic approach to investigate their taxonomic relationships. Phylogenetic analysis of V. splendidus-related strains was carried out using the nucleotide sequences of 16S ribosomal DNA (16S rDNA) and gyrase B subunit (gyrB) genes. Species delineation based on 16S rDNA-sequencing is limited because of divergence between cistrons, roughly equivalent to divergence between strains. Despite a high level of sequence similarity, strains were separated into 2 clades. In the phylogenetic tree constructed on the basis of gyrB gene sequences, strains were separated into 5 independent clusters containing V. splendidus, V. lentus, V. chagasii-type strains and a putative new genomic species. This phylogenetic grouping was almost congruent with that based on DNA-DNA hybridisation analysis. V. pomeroyi, V. kanaloae and V. tasmaniensis-type strains clustered together in a fifth clade. The gyrB gene-sequencing approach is discussed as an alternative for investigating the taxonomy of Vibrio species.


Regulation of transforming growth factor-β1-driven lung fibrosis by galectin-3.
Idiopathic pulmonary fibrosis (IPF) is a chronic dysregulated response to alveolar epithelial injury with differentiation of epithelial cells and fibroblasts into matrix-secreting myofibroblasts resulting in lung scaring. The prognosis is poor and there are no effective therapies or reliable biomarkers. Galectin-3 is a β-galactoside binding lectin that is highly expressed in fibrotic tissue of diverse etiologies. To examine the role of galectin-3 in pulmonary fibrosis. We used genetic deletion and pharmacologic inhibition in well-characterized murine models of lung fibrosis. Further mechanistic studies were performed in vitro and on samples from patients with IPF. Transforming growth factor (TGF)-β and bleomycin-induced lung fibrosis was dramatically reduced in mice deficient in galectin-3, manifest by reduced TGF-β1-induced EMT and myofibroblast activation and collagen production. Galectin-3 reduced phosphorylation and nuclear translocation of β-catenin but had no effect on Smad2/3 phosphorylation. A novel inhibitor of galectin-3, TD139, blocked TGF-β-induced β-catenin activation in vitro and in vivo and attenuated the late-stage progression of lung fibrosis after bleomycin. There was increased expression of galectin-3 in the bronchoalveolar lavage fluid and serum from patients with stable IPF compared with nonspecific interstitial pneumonitis and controls, which rose sharply during an acute exacerbation suggesting that galectin-3 may be a marker of active fibrosis in IPF and that strategies that block galectin-3 may be effective in treating acute fibrotic exacerbations of IPF. This study identifies galectin-3 as an important regulator of lung fibrosis and provides a proof of principle for galectin-3 inhibition as a potential novel therapeutic strategy for IPF.


Influence of heat on biological activity and concentration of oleocanthal--a natural anti-inflammatory agent in virgin olive oil.
The olive oil phenolic oleocanthal is a natural nonsteroidal anti-inflammatory compound that irritates the oral pharynx in a dose-dependent manner. It has been proposed that the biological activity of oleocanthal is partially responsible for the beneficial health effects of the Mediterranean diet. Virgin olive oil containing oleocanthal is often added as an ingredient in a number of cooked dishes, and therefore it is of great importance to understand how best to preserve the putative health-promoting benefits of this compound, as olive oil phenolics are subject to degradation upon heating in general. One extra virgin olive oil containing 53.9 mg/kg oleocanthal was heated at various temperatures (100, 170, and 240 degrees C) for set time periods (0, 1, 5, 20, 60, and 90 min). Oleocanthal concentrations were quantified using HPLC, and its biological activity was determined with a taste bioassay measuring the intensity of throat irritation. Results demonstrated that oleocanthal was heat stable compared with other olive oil phenolics, with a maximum loss of 16% as determined by HPLC analysis. However, there was a significant decrease of up to 31% (p < 0.05) in the biological activity of oleocanthal as determined by the taste bioassay. Although there was minimal degradation of oleocanthal concentration, there was a significant decrease in the biological activity of oleocanthal upon extended heating time, indicating a possible loss of the putative health -benefiting properties of oleocanthal. Alternatively, the difference in the concentration and biological activity of oleocanthal after heat treatment could be a result of an oleocanthal antagonist forming, decreasing or masking the biological activity of oleocanthal.


Accessing timely rehabilitation services for a global aging society? Exploring the realities within Canada's universal health care system.
The proportion of older persons is increasing in developed and developing countries: this aging trend can be viewed as a two-edged sword. On the one hand, it represents remarkable successes regarding advances in health care; and on the other hand, it represents a considerable challenge for health systems to meet growing demand. A growing disequilibrium between supply and demand may be particularly challenging within publicly funding health systems that 'guarantee' services to eligible populations. Rehabilitation, including physical therapy, is a service that if provided in a timely manner, can maximize function and mobility for older persons, which may in turn optimize efficiency and effectiveness of overall health care systems. However, physical therapy services are not considered an insured service under the legislative framework of the Canadian health system, and as such, a complex public/private mix of funding and delivery has emerged. In this article, we explore the consequences of a public/private mix of physical therapy on timely access to services, and use the World Health Organization (WHO) health system performance framework to assess the extent to which the emerging system influences the goal of aggregated and equitable health. Overall, we argue that a shift to a public/private mix may not have positive influences at the population level, and that innovative approaches to deliver services would be desirable to strengthening rather than weaken the publicly funded system. We signal that strategies aimed at scaling up rehabilitation interventions are required in order to improve health outcomes in an evolving global aging society.


Spectrophotometric analysis at the single-cell level: elucidating dispersity within melanic immortalized cell populations.
It is widely held that the melanosome is an exemplar of the absorption features of melanin-containing cells, which are assumed to be uniform in both size and optical characteristics. In recent years, however, it has become increasingly apparent that this is a strikingly poor assumption. Indeed, melanin extracted from natural sources and synthetic melanin both show wide variability in their degree of polymerization (molecular weight) and spectroscopic characteristics. In the current study, imaging spectrophotometry performed on individual cells of immortalized melanin-producing cell lines revealed broad distributions in their sizes: 9.5-36.2 μm for Hs936 human melanoma cells, 10.9-20.8 μm for T47D human breast cancer cells, 5.3-43.5 μm for B16F1 mouse melanoma cells, and 6.4-54.2 μm for B16F10 mouse melanoma cells. The color appearance (from translucent to yellow to nearly black), absorption spectrum, and absorption (extinction) coefficient at 532 nm (28.73 to 364.75, 0.01 to 40.17, 5.88 to 977.19, and 0.01 to 1120 cm-1 for Hs936, T47D, B16F1, and B16F10 cells, respectively) of an individual cell also vary widely and cannot be adequately described by a 'typical' value. In comparison, human red blood cells are much more uniform in size (6.0-8.1 μm diameter; 1.9-3.2 μm thickness), although they too show a broad range of absorptivities, with extinction coefficients in the range of 65 to 370 cm-1 when measured at 532 nm. To further evaluate the impact of these findings on photoacoustic bioanalysis, we performed simulations of the generation of photoacoustic signals expected from these cell types. These simulations revealed that their variation in optical features exerts a pronounced effect on the amplitude and shape of the photoacoustic signals generated from these cell types. Finally, we compared the photoacoustic signal generated from these cells under ideal conditions (i.e., a single cell in isolation) versus a heterogeneous real-world sample, demonstrating that when a single or few cancer cells are present within a blood droplet, the photoacoustic signal is indistinguishable from that measured from blood alone. These outcomes have important ramifications for the early photoacoustic detection of cancer cells and circulating tumor emboli, while pointing to the potential of single-cell imaging spectrophotometry to assess heterogeneity within cell populations in more quantitative terms.


Disrupted brain network topology in Parkinson's disease: a longitudinal magnetoencephalography study.
Although alterations in resting-state functional connectivity between brain regions have previously been reported in Parkinson's disease, the spatial organization of these changes remains largely unknown. Here, we longitudinally studied brain network topology in Parkinson's disease in relation to clinical measures of disease progression, using magnetoencephalography and concepts from graph theory. We characterized whole-brain functional networks by means of a standard graph analysis approach, measuring clustering coefficient and shortest path length, as well as the construction of a minimum spanning tree, a novel approach that allows a unique and unbiased characterization of brain networks. We observed that brain networks in early stage untreated patients displayed lower local clustering with preserved path length in the delta frequency band in comparison to controls. Longitudinal analysis over a 4-year period in a larger group of patients showed a progressive decrease in local clustering in multiple frequency bands together with a decrease in path length in the alpha2 frequency band. In addition, minimum spanning tree analysis revealed a decentralized and less integrated network configuration in early stage, untreated Parkinson's disease that also progressed over time. Moreover, the longitudinal changes in network topology identified with both techniques were associated with deteriorating motor function and cognitive performance. Our results indicate that impaired local efficiency and network decentralization are very early features of Parkinson's disease that continue to progress over time, together with reductions in global efficiency. As these network changes appear to reflect clinically relevant phenomena, they hold promise as markers of disease progression.


Risk factors for bacteriological quality of bulk tank milk in Prince Edward Island dairy herds. Part 1: overall risk factors.
The objective of this study was to determine on-farm risk factors for bacteriological quality of bulk tank milk. Bulk tank raw milk quality was evaluated on all Prince Edward Island dairy herds (n = 235) over a 2-yr period (March 2005 to March 2007). Biweekly total bacterial, preliminary incubation, laboratory pasteurization, and coliform counts were conducted using a Petrifilm culture system. For the assessment of risk factors, a case-control study was conducted from January 2006 to May 2007. Case and control herds were defined based on the last 6 analyses of bulk tank bacterial counts before on-farm evaluation. Cases were herds that had multiple elevated counts for any of the parameters measured. A total of 69 herds (39 cases and 30 control herds) were evaluated. Data collection included 1) observation and questionnaire on basic hygiene and farm management practices; 2) complete wash analysis of the milking equipment, monitoring the presence of bacterial films on equipment and evaluation of cooling system function; and 3) environmental and cow hygiene scoring. Data were analyzed using multivariable logistic regression. The results of the final model indicated that high alkalinity in the wash water and poor teat-end cleanliness were associated with high bacterial counts in bulk tank milk (odds ratios = 12 and 5.3, respectively). It was also observed that high water temperature of detergent wash and the use of a water softener were associated with low bacterial counts in bulk tank milk (odds ratios = 0.87 and 0.11, respectively). A significant association between udder hair clipping and teat-end cleanliness was also observed. In conclusion, this study highlights the importance of udder hygiene and milking system washing factors on hygienic quality of bulk tank milk.


Functional CD40 ligand expressed by human blood dendritic cells is up-regulated by CD40 ligation.
We examined whether human blood dendritic cells (DCs) express a functional ligand for CD40 (CD40L). Human blood DCs expressed significant amounts of cell surface CD40L identical to that expressed on activated T cells, as detected by mAb to CD40L or a chimeric CD40.Ig fusion protein (CD40.Ig). Stimulation through CD40 up-regulated protein and mRNA CD40L expression in DCs, B cells, and B cell lines. CD40-mediated CD40L expression was inhibited by a protein tyrosine kinase inhibitor, herbimycin, in a dose-dependent manner, suggesting that the induction of CD40L expression via CD40 requires protein tyrosine kinase activity. CD40L surface expression correlated with constitutive or inducible levels of CD40L-specific mRNA, as determined by reverse transcribed PCR analysis (RT-PCR) using CD40L-homologous primers. Furthermore, CD40L on DCs was functional, since CD40L+ DCs, unlike CD40L- DCs, induced B cell IgG and IgA production, and this induction could be inhibited by blocking CD40L-CD40 interactions with mAb to CD40L. Thus, CD40L on DCs and CD40L induced by crosslinking CD40 may regulate B cell activation and maturation. The importance of DC CD40L expression on B cell function is discussed.


Muscle weakness is a prognostic indicator of disability and chronic disease multimorbidity.
The objective of this study was to use nationally-representative data on Americans greater than 50 years of age to determine the association between grip strength and inflammation as independent predictors of incident disability, chronic multimorbidity and dementia. Middle age and older adults (n = 12,618) from the 2006-2008 waves of the Health and Retirement Study with 8-years of follow-up were included. Longitudinal modeling was performed to examine the association between baseline grip strength (normalized to body mass: NGS) and high sensitivity C-reactive protein (hs-CRP) (≥3.0 mg/L) with incident physical disabilities (i.e., ≥2 limitations to activities of daily living), chronic multimorbidity (≥2 of chronic conditions), and dementia. The odds of incident disability were 1.25 (95% CI: 1.20-1.30) and 1.31 (95% CI: 1.26-1.36) for men and women respectively, for each 0.05-unit lower NGS. The odds of incident chronic multimorbidity were 1.14 (95% CI: 1.08-1.20) and 1.14 (95% CI: 1.07-1.21) for men and women respectively for each 0.05-unit lower NGS. The odds of incident dementia were 1.10 for men (95% CI: 1.02-1.20) for each 0.05-unit lower NGS, but there was no significant association for women. Elevated hs-CRP was only associated with chronic multimorbidity among men (OR = 1.29; 95% CI: 1.00-1.73) and women (OR = 1.60; 95% CI: 1.26-2.02). Our findings indicate a robust inverse association between NGS and disability and chronic, multimorbidity in older men and women, and dementia in men. Elevated hs-CRP was only associated with chronic multimorbidity in men and women. Healthcare providers should implement measures of grip strength in routine health assessments and discuss the potential dangers of weakness as well as interventions to improve strength with their patients.


Novel Use of Responsive Neurostimulation (RNS System) in the Treatment of Super Refractory Status Epilepticus.
There are very few randomized controlled trials studying treatment of super refractory status epilepticus (SE), despite estimated occurrence in about 15% of SE cases and its association with high morbidity and mortality rates. Small case series and case reports have described use of neurostimulation, including vagal nerve stimulation, transcranial magnetic stimulation, and deep brain stimulation, to treat super refractory SE when medical interventions have failed. To our knowledge, this is the first reported case of responsive neurostimulation being used to successfully treat a case of super refractory SE. A 37-year-old man with refractory focal epilepsy and a known focal cortical dysplasia involving motor cortex was implanted with an RNS System device after being in super refractory SE for 20 days. Responsive neurostimulation strip and depth electrodes were placed targeting the cortical dysplasia. Detection and stimulation parameters were adjusted over a 14-day period, as medications were gradually weaned. Seizures abated 15 days after implant, 24 hours after stimulation parameters were configured to mimic seizure offset pattern. Seizure remission was sustained, allowing the patient to be weaned off sedating medications and discharged to a rehabilitation facility. At 6 weeks of follow-up, the patient was near his neurologic baseline with no focal deficits.


Swine flu and antibiotics.
Health services worldwide are likely to be hard-pressed by swine flu-related illness in the months ahead. Secondary infections with Streptococcus pneumoniae, other streptococci (e.g. Streptococcus pyogenes), Haemophilus influenzae and Staphylococcus aureus are likely to be important causes of morbidity and mortality. The UK Department of Health recently published clinical pathways for the management of swine flu. Suggested severity criteria have not been validated in respiratory infection and are different from those previously published. Antibiotics are recommended for all patients assessed at hospital, regardless of severity of illness; cephalosporins or quinolones are suggested for inpatients with pneumonia. These recommendations will jeopardize recent decreases in Clostridium difficile-associated diarrhoea (CDAD) and methicillin-resistant S. aureus (MRSA) in UK hospitals. This article, written on behalf of the BSAC Council, considers these recommendations and provides alternative antibiotic regimens for a range of clinical scenarios.


What determines motor neuron number? Slow scaling of facial motor neuron numbers with body mass in marsupials and primates.
How does the number of motor neurons in the brain correlate with the muscle mass to be controlled in the body? Numbers of motor neurons are known to be adjusted during development by cell death, but the change in the percentage of surviving motor neurons in response to experimental changes in target muscle mass is relatively small. Here we address the quantitative matching between final numbers of motor neurons in the facial nucleus and body mass (which we use as a proxy for the muscle mass). In 22 marsupial species, we found that the number of facial motor neurons is strongly correlated with body mass, and scales across species as a power function of body mass with a very small exponent of 0.184, which is close to the exponent found in primates from previously published data. With such an exponent, doubling the body mass is accompanied by a modest increase of only 14% in numbers of facial motor neurons, while halving body mass results in a decrease of only 12%. These numbers are remarkably similar to the 15-20% increase or 8% decrease in the number of spinal cord motor neurons that results from experimental or natural doubling or reducing by half the target muscle field of birds and amphibians. The scaling rule presented here might thus account for the quantitative matching of motor neurons to their target muscle mass in evolution. With this small scaling exponent, our data also raise the possibility that larger animals will have larger motor units.


Knee moment and shear force are correlated with femoral tunnel orientation after single-bundle anterior cruciate ligament reconstruction.
Increasing evidence has shown that anatomic single-bundle anterior cruciate ligament reconstruction (ACLR) better restores normal knee kinematics and functionality than nonanatomic ACLR. Whether anatomic reconstruction results in better knee kinetics during daily activities has not been fully investigated. To assess the relationship between femoral tunnel angle and kinetic parameters of the knee joint during walking after single-bundle ACLR and to compare the radiographic and kinetic results of patients who underwent anatomic ACLR with those of patients who underwent nonanatomic ACLR. Controlled laboratory study. Twenty-one patients who underwent unilateral ACLR were recruited, and 20 healthy subjects from a previous study were used as a control group. All surgical procedures were performed by a single surgeon, 11 using the transtibial (TT) technique and 10 using the anteromedial portal (AMP) technique. Femoral tunnel orientation was measured from posterior-to-anterior radiographs. Dynamic knee joint moments and shear forces during gait were evaluated using 3-dimensional motion analysis and inverse dynamics. Relationships between femoral tunnel angles and kinetic results were evaluated via linear regression. Results were compared between 2 ACLR groups and controls using 1-way analysis of variance. Femoral tunnel angle had significant correlations with peak external knee flexion moment and posterior shear force during early stance. The TT group had a significantly smaller (more vertical) mean femoral tunnel angle (19.4° ± 4.1°) than the AMP group (36.4° ± 5.8°). Significant reductions were found in the normalized peak external knee flexion moment (TT, 0.15 ± 0.12 Nm/kg·m; AMP, 0.25 ± 0.12 Nm/kg·m; control, 0.25 ± 0.16 Nm/kg·m) (P = .032) and posterior shear force (TT, 0.64 ± 0.55 N/kg; AMP, 1.10 ± 0.58 N/kg; control, 1.35 ± 0.55 N/kg) (P = .024) in the TT group compared with controls, but not in the AMP group. Moreover, a significantly greater medial shear force was found in the TT group during the late stance phase (TT, 1.08 ± 0.32 N/kg; AMP, 0.89 ± 0.26 N/kg; control, 0.83 ± 0.22 N/kg) (P = .038). A greater peak external knee adduction moment was found in both ACL groups during the early stance phase (TT, 0.25 ± 0.07 Nm/kg·m; AMP, 0.25 ± 0.07 Nm/kg·m; control, 0.19 ± 0.05 Nm/kg·m) (P < .01). Knee joint kinetic changes are seen within months (~10 months) after ACLR. This study revealed significant relationships between femoral tunnel orientation and postoperative knee joint flexion moment and posterior shear force during walking. The AMP technique provides better restoration of these knee kinetic parameters compared with the TT technique at this postoperative time point. The femoral tunnel angle measured from plain radiographs can be used as an important metric of postoperative knee joint kinetics. This information provides a better understanding of the knee joint's biomechanical environment after ACLR using commonly used single-bundle techniques.


Optically-isotropic nanoencapsulated liquid crystal displays based on Kerr effect.
An innovative liquid crystal display (LCD), incorporating in-plane switching mechanism with LC nanoencapsulation, is developed and its electro-optic (EO) performances are characterized. The nanoencapsulated LCDs exhibit electric-field induced transitions from an optically-isotropic state to an anisotropic one based on Kerr effect. Extremely small size of LC droplets embedded in the nanoencapsulated layer, less than a quarter of wavelength of incident light, appear transparent in the voltage-off state and gradually becomes birefringent in response to applied voltage, exhibiting the optical behavior of uniaxial retarder. Due to optically-isotropic nature and a large Kerr effect, they show wide viewing angle characteristics and good EO performances.


Melioidosis in Thailand: Present and Future.
A recent modelling study estimated that there are 2800 deaths due to melioidosis in Thailand yearly. The Thailand Melioidosis Network (formed in 2012) has been working closely with the Ministry of Public Health (MoPH) to investigate and reduce the burden of this disease. Based on updated data, the incidence of melioidosis is still high in Northeast Thailand. More than 2000 culture-confirmed cases of melioidosis are diagnosed in general hospitals with microbiology laboratories in this region each year. The mortality rate is around 35%. Melioidosis is endemic throughout Thailand, but it is still not uncommon that microbiological facilities misidentify Burkholderia pseudomallei as a contaminant or another organism. Disease awareness is low, and people in rural areas neither wear boots nor boil water before drinking to protect themselves from acquiring B. pseudomallei. Previously, about 10 melioidosis deaths were formally reported to the National Notifiable Disease Surveillance System (Report 506) each year, thus limiting priority setting by the MoPH. In 2015, the formally reported number of melioidosis deaths rose to 112, solely because Sunpasithiprasong Hospital, Ubon Ratchathani province, reported its own data (n = 107). Melioidosis is truly an important cause of death in Thailand, and currently reported cases (Report 506) and cases diagnosed at research centers reflect the tip of the iceberg. Laboratory training and communication between clinicians and laboratory personnel are required to improve diagnosis and treatment of melioidosis countrywide. Implementation of rapid diagnostic tests, such as a lateral flow antigen detection assay, with high accuracy even in melioidosis-endemic countries such as Thailand, is critically needed. Reporting of all culture-confirmed melioidosis cases from every hospital with a microbiology laboratory, together with final outcome data, is mandated under the Communicable Diseases Act B.E.2558. By enforcing this legislation, the MoPH could raise the priority of this disease, and should consider implementing a campaign to raise awareness and melioidosis prevention countrywide.


Phylogenetic characterization of epibiotic bacteria in the accessory nidamental gland and egg capsules of the squid Loligo pealei (Cephalopoda:Loliginidae).
Sexually mature female squid Loligo pealei harbour dense bacterial communities in their accessory nidamental glands (ANGs) and in their egg capsules (ECs). This study describes a molecular approach using the 16S rRNA gene (rDNA) to identify bacterial populations within the ANG and the ECs of the North Atlantic squid species L. pealei. Fluorescent in situ hybridization (FISH) and 16S rDNA analysis showed that predominantly alpha- and, to a lesser extent, gamma-proteobacteria were the predominant components of the ANG and EC bacterial communities. Sequencing results showed the presence of alpha-proteobacterial populations affiliated with the Roseobacter group and additional deep-branching alpha-proteobacterial lineages. In contrast, isolates from the ANG and ECs contained only a few alpha-proteobacteria of the Roseobacter group compared with several gamma-proteobacterial isolates, mostly Shewanella and Pseudoalteromonas species. Most of the ANG-associated bacterial populations were also found within the ECs of L. pealei. The molecular approach allowed the visualization of alpha-proteobacteria as major constituents of a bacterial symbiosis within the reproductive system of the Loliginidae.


Neuromuscular Exercises Improve Shoulder Function More Than Standard Care Exercises in Patients With a Traumatic Anterior Shoulder Dislocation: A Randomized Controlled Trial.
There is an important gap in knowledge about the effectiveness of nonoperative treatment (exercise) for patients with traumatic primary and recurrent anterior shoulder dislocations (ASDs). The purpose of this study was to assess the efficacy and safety of physical therapist-supervised, shoulder instability neuromuscular exercise (SINEX) versus self-managed, home-based, standard care shoulder exercise (HOMEX) in patients with traumatic ASDs. The hypothesis was that SINEX would have a larger effect and fewer adverse events compared with HOMEX. Randomized controlled trial; Level of evidence, 2. A total of 56 participants with radiographically verified, trauma-initiated primary or recurrent ASDs and self-reported decreased shoulder function were randomized to 12 weeks of either SINEX or HOMEX. The SINEX program consisted of 7 exercises, individually progressing from basic (2 × 20 repetitions each day) to elite (2 × 10 repetitions, 3 times weekly). The HOMEX program included 5 shoulder exercises performed 3 times weekly (2 × 10 repetitions). The primary outcome was the Western Ontario Shoulder Instability Index (WOSI) score, ranging from 0 (best possible) to 2100. The between-group minimal clinically important difference at 12 weeks was 250 points. Secondary outcomes included WOSI subdomain scores, patient-reported ratings of kinesiophobia and pain, objective shoulder function, patient satisfaction, and number of adverse events. The between-group mean difference in the WOSI total score at 12 weeks significantly favored SINEX over HOMEX (-228.1 [95% CI, -430.5 to -25.6]). SINEX was furthermore superior to HOMEX in most of the secondary outcomes (3/4 subdomains of the WOSI and pain level during the past 7 days as well as clinical signs of anterior shoulder instability). Also, although not statistically significant, less than half the proportion of the SINEX patients compared with the HOMEX patients (3/27 [11%] vs 6/24 [25%], respectively; P = .204) underwent or were referred for shoulder stabilizing surgery. Satisfaction with both exercise programs was high, and no serious adverse events were reported. Neuromuscular shoulder exercise (SINEX) was superior to standard care exercise (HOMEX) in patients with traumatic ASDs. Further long-term follow-ups on treatment effects are needed. NCT02371928 (ClinicalTrials.gov identifier).


Insights into the role of the junctional region of Plasmodium falciparum dihydrofolate reductase-thymidylate synthase.
Plasmodium falciparum dihydrofolate reductase-thymidylate synthase (pfDHFR-TS) is a well-defined target of anti-malarial drug, such as pyrimethamine and cycloguanil. Emergence of malaria parasites resistant to these drugs has been shown to be associated with point mutations of the gene coding for the target enzymes. Although the 3D-structure of P. falciparum bifunctional pfDHFR-TS has been reported previously, relatively little is known about the interactions between the pfDHFR and pfTS domains and the roles of the junctional region that links the two domains together. Therefore, a thorough understanding of the interaction of the two domains and the role of the junctional region of this target is important as the knowledge could assist the development of new effective anti-malarial drugs aimed at overcoming drug-resistant malaria. A system was developed to investigate the interaction between pfDHFR and pfTS domains and the role of the junctional region on the activity of the recombinant pfTS. Based on the ability of co-transformed plasmids coding for pfDHFR and pfTS with truncated junctional region to complement the growth of TS-deficient Escherichia coli strain χ2913recA(DE3) on minimum media without thymidine supplementation, active pfTS mutants with minimal length of junctional region were identified. Interactions between active pfDHFR and the pfTS domains were demonstrated by using a bacterial two-hybrid system. Using TS-deficient E. coli strain χ2913recA(DE3), the authors have shown for the first time that in P. falciparum a junctional region of at least 44 amino acids or longer was necessary for the pfTS domain to be active for the synthesis of thymidylate for the cells. Truncation of the junctional region of the bifunctional pfDHFR-TS further confirmed the above results, and suggested that a critical length of the junctional peptide of pfDHFR-TS would be essential for the activity of TS to catalyze the synthesis of thymidylate. The present study demonstrated the interactions between the pfDHFR and pfTS domains of the bifunctional pfDHFR-TS, and revealed that the junctional region linking the two protein domains is essential for the expression of catalytically active pfTS domain. The findings could be useful since inhibition of the pfDHFR-TS domain-domain interaction could form a basis for the development of new anti-malarial drugs based on targeting the non-active site region of this important enzyme.


Perindopril-based blood pressure-lowering therapy reduces amino-terminal-pro-B-type natriuretic peptide in individuals with cerebrovascular disease.
The plasma amino-terminal-pro-B-type natriuretic peptide (NT-proBNP) level predicted congestive heart failure, myocardial infarction, and ischaemic stroke in participants of the Perindopril Protection Against Recurrent Stroke Study (PROGRESS), a placebo-controlled study of the effects of blood pressure lowering on cardiovascular events among individuals with cerebrovascular disease. Active treatment comprised a flexible regimen based on perindopril, with the addition of indapamide at the discretion of treating physicians. Active treatment reduced cardiovascular events, and we therefore investigated whether active treatment modified NT-proBNP and other cardiovascular risk factors. We measured NT-proBNP and other cardiovascular risk factors at randomization and after 13 months of therapy in a subset of 357 PROGRESS participants. Baseline systolic and pulse pressures were higher in individuals with elevated baseline NT-proBNP levels. In comparison with placebo, active treatment reduced the blood pressure and NT-proBNP levels, and increased renin levels. Reduction of NT-proBNP levels by active treatment was most evident in individuals with baseline NT-proBNP levels in the highest quarter (> 26 pmol/l), with a median reduction of 16 pmol/l (interquartile range 0-51 pmol/l, P = 0.004), corresponding to a median decrease of 39% (interquartile range 0-69%). Active treatment reduced blood pressure similarly for individuals in each of the four quarters of baseline NT-proBNP. Active therapy had no effect on plasma lipid, C-reactive protein, homocysteine, or soluble vascular cell adhesion molecule 1 levels. We conclude that plasma NT-proBNP level, in addition to predicting cardiovascular risk, may provide a measure of risk reduction by blood pressure-lowering therapy.


Excited state C-N bond dissociation and cyclization of tri-aryl amine-based OLED materials: a theoretical investigation.
Tri-aryl amines (TAA) such as triphenylamine (TPA) are widely used for designing chromophores for dye-sensitized solar cells (DSSC) and organic light-emitting diodes (OLED). These materials degrade over time and hence result in reduced performance. Therefore, exploring the associated mechanistic pathways and factors controlling the degradation is necessary for future development of durable TAA-based devices. Hence, in this study, the complete active space self-consistent-field (CASSCF) method coupled with second order N-electron valence perturbation theory (NEVPT2) calculations was carried out to understand the excited state phenomena occurring in TAA using TPA and N,N-diphenyl-2-naphthylamine (DPNA) as model systems. The results indicated the presence of a conical intersection between ground and first excited singlet states with C-N bond dissociation, which acts as a channel for the excited molecules to dissociate and form radical fragments (phenyl/naphthyl). This occurrence is unusual for non-saturated bonds with delocalization. The resulting radical fragments formed intra-molecular products and subsequently yielded five- and six-membered cyclized products depending on the type of aryl groups. The significant findings from this study throw light on the photostability of TAA-based OLED devices as well as on the possible route to synthesize cyclic amines such as carbazoles.


Simulation training improves clinical knowledge of major haemorrhage management in foundation year doctors.
(i) To develop a major haemorrhage simulation training programme. (ii) To design an assessment tool to measure the effectiveness of this programme. (iii) To use simulation training to create more effective protocols. Major haemorrhage is a time-critical medical emergency that can be faced by every Foundation Year (FY) doctor. Standard methods of teaching provide limited opportunity for junior doctors to improve their knowledge and practical skills for major haemorrhage situations. Simulation is increasingly used in medical training but has not been used as a means both to facilitate learning and refine hospital major haemorrhage policy. The effect of major haemorrhage simulation on attendees' learning was compared to a comparator group not exposed to simulation. Questionnaire pre-simulation and 3 months post-simulation training assessed knowledge of the Trust Major Haemorrhage Protocol (MHP). Sixteen FY1 doctors attended simulation training. The comparator group included 47 FY1 doctors. No significant difference was found between simulation and comparator groups on baseline questionnaire scores. The simulation group showed significant improvement (total score, mean standard deviation (SD) pre-test 27.15 (4.02), post-test 39.13 (3.91), p < 0.001). The comparator group showed no significant change (total score, pre-test 25.06 (5.70), post-test 23.54 (6.85), p = 0.33) (19 lost to follow up). The study resulted in the production of a new MHP. This study has demonstrated that simulation training improved doctors' knowledge in major haemorrhage management and that the experience of observing the simulation training allowed senior staff to undertake analysis and improvement of an existing MHP.


Stability of polymer glasses vitrified under stress.
How stress or strain imparts mobility to glasses is a scientific issue linking ideas of jamming and the glass transition across colloids, granular materials, polymers, and molecular glasses. Here, we address for the first time how stress applied during vitrification, formation of the glassy state by a temperature quench, affects the subsequent stability of the glassy state, even after the stress has been removed. Using entangled polymers that are easily manipulated mechanically above the glass transition temperature, we find that the resulting polymer glasses become less stable, exhibiting a higher physical aging rate, when stress is applied while rapidly cooling the polymer films. The data show an initial plateau value at low stress, before transitioning rapidly to a higher aging rate at larger stress. These results are suggestive of the glassy system being left trapped in a less stable, higher energy state with faster physical aging rate when stressed above some minimum value during vitrification.


Age-related deregulation of TDP-43 after stroke enhances NF-κB-mediated inflammation and neuronal damage.
TDP-43 has been identified as a disease-associated protein in several chronic neurodegenerative disorders and increasing evidence suggests its potentially pathogenic role following brain injuries. Normally expressed in nucleus, under pathological conditions TDP-43 forms cytoplasmic ubiquitinated inclusions in which it is abnormally phosphorylated and cleaved to generate a 35 and a 25 kDa C-terminal fragments. In the present study, we investigated age-related expression patterns of TDP-43 in neurons and glia and its role as modulator of inflammation following ischemic injury. Wild-type and TDP-43 transgenic mice of different age groups were subjected to transient middle cerebral artery occlusion. The role of TDP-43 in modulation of inflammation was assessed using immunofluorescence, Western blot analysis, and in vivo bioluminescence imaging. Finally, post-mortem stroke human brain sections were analyzed for TDP-43 protein by immunohistochemistry. We report here an age-related increase and formation of ubiquitinated TDP-43 cytoplasmic inclusions after stroke. The observed deregulation in TDP-43 expression patterns was associated with an increase in microglial activation and innate immune signaling as revealed by in vivo bioluminescence imaging and immunofluorescence analysis. The presence of ubiquitinated TDP-43 aggregates and its cleaved TDP-35 and TDP-25 fragments was markedly increased in older, 12-month-old mice leading to larger infarctions and a significant increase in in neuronal death. Importantly, unlike the hallmark neuropathological features associated with chronic neurodegenerative disorders, the TDP-43-positive cytoplasmic inclusions detected after stroke were not phosphorylated. Next, we showed that an increase and/or overexpression of the cytoplasmic TDP-43 drives the pathogenic NF-κB response and further increases levels of pro-inflammatory markers and ischemic injury after stroke in age-dependent manner. Finally, analyses of the post-mortem stroke brain tissues revealed the presence of the cytoplasmic TDP-43 immunoreactive structures after human stroke. Together, our findings suggest that the level of cytoplasmic TDP-43 increases with aging and may act as an age-related mediator of inflammation and neuronal injury after stroke. Thus, targeting cytoplasmic TDP-43 may have a therapeutic potential after stroke.


Social Class Predicts Emotion Perception and Perspective-Taking Performance in Adults.
"Theory of Mind" (ToM; people's ability to infer and use information about others' mental states) varies across cultures. In four studies (N = 881), including two preregistered replications, we show that social class predicts performance on ToM tasks. In Studies 1A and 1B, we provide new evidence for a relationship between social class and emotion perception: Higher-class individuals performed more poorly than their lower-class counterparts on the Reading the Mind in the Eyes Test, which has participants infer the emotional states of targets from images of their eyes. In Studies 2A and 2B, we provide the first evidence that social class predicts visual perspective taking: Higher-class individuals made more errors than lower-class individuals in the Director Task, which requires participants to assume the visual perspective of another person. Potential mechanisms linking social class to performance in different ToM domains, as well as implications for deficiency-centered perspectives on low social class, are discussed.


Community composition of the Planctomycetes associated with different macroalgae.
Insights into the diversity of marine natural microbial biofilms, as for example those developing at the surface of marine macroalgae, can be obtained by using molecular techniques based on 16S rRNA genes. We applied denaturing gradient gel electrophoresis (DGGE) with 16S rRNA genes-specific primers for Planctomycetes to compare the communities of these organisms developing on six different macroalgae (Chondrus crispus, Fucus spiralis, Mastocarpus stellatus, Porphyra dioica, Sargassum muticum, and Ulva sp.) sampled in spring 2012 in two rocky beaches in the north of Portugal. Planctomycetes can be one of the dominant organisms found in the epibacterial community of macroalgae, and we wanted to determine the degree of specificity and the spatial variation of these group. Shannon diversity indexes obtained from the comparison of DGGE profiles were similar in all the macroalgae, and in both sites, F. spiralis was the algae presenting lower Planctomycetes diversity, while M. stellatus and P. dioica from Porto showed the highest diversity. The analysis of DGGE profiles, including anosim statistics, indicate the existence of a specific Planctomycetes community associated with the algal host, likely independent of geographical variation. Sequencing of DGGE bands indicated that Planctomycetes communities were highly diverse, and some Operational Taxonomic Units seemed to be specifically associated with each macroalgae.


Research on evidence-based practices: future directions in an era of recovery.
Many mental health systems are trying to promote the adoption of what has come to be known as evidence-based practices while incorporating a recovery vision into the services they provide. Unfortunately, much of the existing, published, research on evidence-based practices was conceived without an understanding of the recovery vision and/or implemented prior to the emergence of the recovery vision. As result, evidence-based practice research that has been published to date is deficient in speaking to a system being built on a recovery philosophy and mission; these deficiencies are detailed, and suggestions are advanced for new directions in evidence-based practice research.


Identity and function of a large gene network underlying mutagenic repair of DNA breaks.
Mechanisms of DNA repair and mutagenesis are defined on the basis of relatively few proteins acting on DNA, yet the identities and functions of all proteins required are unknown. Here, we identify the network that underlies mutagenic repair of DNA breaks in stressed Escherichia coli and define functions for much of it. Using a comprehensive screen, we identified a network of ≥93 genes that function in mutation. Most operate upstream of activation of three required stress responses (RpoS, RpoE, and SOS, key network hubs), apparently sensing stress. The results reveal how a network integrates mutagenic repair into the biology of the cell, show specific pathways of environmental sensing, demonstrate the centrality of stress responses, and imply that these responses are attractive as potential drug targets for blocking the evolution of pathogens.


Characterization of human autoantibodies that selectively precipitate the 7SL RNA component of the signal recognition particle.
The signal recognition particle (SRP), which consists of the 7SL RNA molecule associated with six polypeptides ranging between 9,000 and 72,000 m.w., mediates the translocation of newly synthesized proteins across the endoplasmic reticulum. We have characterized autoantibodies that are directed against this particle from two patients with rheumatic diseases. These sera immunoprecipitated the 7SL RNA from whole extracts of HeLa cells radiolabeled with 32P, but no RNA from deproteinized cell extracts. From 35S-methionine-labeled cell extracts, they immunoprecipitated a single polypeptide of 54,000 m.w. that is consistent with a known SRP component. Sucrose density gradient studies confirmed that this protein co-migrated with the 7SL RNA, indicating the likelihood that it is physically associated with this RNA. Thus, the 54,000 m.w. SRP protein, which is essential for the SRP functions of elongation arrest and translocation, appears to be a preferential target for human autoimmune responses. Human autoantibodies that recognize the SRP should be useful adjuncts to animal antisera for studies of the structure and function of this particle.


The Achilles Functional Index.
The literature regarding the management of spontaneous rupture of the Achilles tendon is controversial and confusing. The relative infrequency of the condition in any one center prohibits the completion of well-designed clinical studies. Many of the disputes could be addressed and innovations tested if an appropriate animal model were available. We present a method for evaluating Achilles tendon function from measurements of the prints, preserved in bromphenol-blue-impregnated photocopying paper, of the hindfeet of walking rats. The stimulus for this study was derived from de Medinaceli's method for assessing the functional condition of rat sciatic nerves (de Medinaceli L, Freed WJ, Wyatt RJ: An index of the functional condition of rat sciatic nerve based on measurements made from walking tracks. Exp Neurol 77:634-643, 1982). Four variables were measured from these walking tracks, and comparisons between the damaged (experimental) and intact (normal) side were converted to proportional deficits. The relative contribution of each parameter to the overall deficit was determined by multiple linear regression analysis, and the variables were weighted accordingly to obtain an "Achilles Functional Index" (AFI). A sham operation produced no functional deficit, whereas animals subjected to a 0.5-cm midsubstance Achilles tendon defect demonstrated a markedly impaired AFI. Animals with repaired transected Achilles tendons also demonstrated a significant, but less severely impaired AFI. The functional deficit in this repair group returned to control values by postoperative day 15, whereas animals with a defect remained impaired at day 15. Furthermore, an excellent correlation was found between the functional recovery and biomechanical properties (ultimate failure load) of the healing tendon (r = 0.94; p less than 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)


Difference gel electrophoresis.
Difference gel electrophoresis (DIGE) was invented to circumvent the inherent variability of 2-DE. This variability is a natural consequence of separating thousands of proteins over a large space, such as a 15 x 20 cm slab of polyacrylamide gel. The originators of 2-DE envisioned being able to compare cancerous cells and normal cells to understand what makes these cells different. Gel-to-gel variability made this an extremely difficult task. We reasoned that if both samples could be run on the same gel, then the inherent variability would be obviated. Thus, we created matched sets of fluorescent dyes that allows one to compare two or three protein samples on a single gel. In the 12 years since the description of DIGE first appeared in Electrophoresis, this founding paper has been cited over 660 times. This review highlights some of the improvements and applications of DIGE. We hope these examples are illustrative of what has been done and where the field is headed.


Effects of supplementing low-protein diets with sodium dichloroacetate and glucose on growth performance, carcass traits, and meat quality of growing-finishing pigs.
The objective of this study was to evaluate the effects of supplementing low-protein diets with sodium dichloroacetate (DCA) and glucose on growth performance, carcass traits, and meat quality of growing-finishing pigs. A total of 80 crossbred (Duroc × Landrace × Large White) growing barrows (27 ± 0.4 kg body weight) were allocated randomly to one of the five treatments during three successive 4-wk periods. There were five diets in each phase. Diet 1 was the control diet with normal protein levels (CON) where protein levels in the three phases were 18%, 16.5%, and 15.5%, respectively. The dietary protein levels of Diets 2, 3, 4, and 5 (the low-protein diets, LP) were decreased by 4.5% compared to Diet 1. Additionally, Diets 3 and 4 were supplemented with an extra 120 mg/kg DCA (LP + DCA) or 1.8% glucose (LP + GLUC), respectively. Diet 5 was further supplemented with an extra 120 mg/kg DCA and 1.8% glucose (LP + DCA + GLUC). The LP + DCA diet increased the average daily weight gain of pigs compared to the CON and LP diet in phase 3 and the overall experimental period (P < 0.001). The LP diet reduced the gain:feed ratios of the pigs compared to the CON, LP + DCA, and LP + DCA + GLUC diets in phase 1 and the overall experimental period (P < 0.001). Furthermore, gain:feed ratios in LP + DCA and LP + DCA + GLUC groups did not differ from that of the CON group (P > 0.10). Pigs fed the LP + DCA diet had higher pH values of meat at 24 h post-mortem than the CON group (P < 0.05). The LP + DCA + GLUC diet increased the total protein content in the longissimus dorsi (LD) muscle of pigs, compared to the other dietary treatments (P < 0.05), and increased the Arg and Leu contents in the LD muscle compared to the LP + DCA diet (P < 0.05). Moreover, the LP + DCA diet induced a higher C18:1n9t percentage in the LD muscle of pigs compared to other groups (P < 0.05). In conclusion, an LP diet reduced the feed efficiency in pigs and barely affected meat quality, whereas 120 mg/kg DCA supplementation in an LP diet improved the growth performance of growing-finishing pigs, showed modest effects on carcass traits, and improved the muscle protein content with the addition of glucose.


Development of modularity in the neural activity of children's brains.
We study how modularity of the human brain changes as children develop into adults. Theory suggests that modularity can enhance the response function of a networked system subject to changing external stimuli. Thus, greater cognitive performance might be achieved for more modular neural activity, and modularity might likely increase as children develop. The value of modularity calculated from functional magnetic resonance imaging (fMRI) data is observed to increase during childhood development and peak in young adulthood. Head motion is deconvolved from the fMRI data, and it is shown that the dependence of modularity on age is independent of the magnitude of head motion. A model is presented to illustrate how modularity can provide greater cognitive performance at short times, i.e. task switching. A fitness function is extracted from the model. Quasispecies theory is used to predict how the average modularity evolves with age, illustrating the increase of modularity during development from children to adults that arises from selection for rapid cognitive function in young adults. Experiments exploring the effect of modularity on cognitive performance are suggested. Modularity may be a potential biomarker for injury, rehabilitation, or disease.


The Effect of Perioperative Vitamin C on Postoperative Analgesic Consumption: A Meta-Analysis of Randomized Controlled Trials.
Because the analgesic effect of vitamin C against acute pain remains poorly addressed, this meta-analysis aimed at investigating its effectiveness against acute postoperative pain. A total of seven randomized controlled trials with placebo/normal controls were identified from PubMed, Cochrane Library, Medline, Google Scholar, and Embase databases. Pooled analysis showed a lower pain score (standardized mean difference (SMD) = -0.68, 95% CI: -1.01 to -0.36, p < 0.0001; I2 = 57%) and a lower morphine consumption (weighted mean difference (WMD) = -2.44 mg, 95% CI: -4.03 to -0.86, p = 0.003; I2 = 52%) in the vitamin group than that in the placebo group within postoperative 1-2 h. At postoperative 24 h, a lower pain score (SMD = -0.65, 95% CI: -1.11 to -0.19, p = 0.005; I2 = 81%) and lower morphine consumption (WMD = -6.74 mg, 95% CI: -9.63 to -3.84, p < 0.00001; I2 = 85%) were also noted in the vitamin group. Subgroup analyses demonstrated significant reductions in pain severity and morphine requirement immediately (1-2 h) and 24 h after surgery for patients receiving intravenous vitamin C but not in the oral subgroup. These findings showed significant reductions in pain score and opioid requirement up to postoperative 24 h, respectively, suggesting the effectiveness of perioperative vitamin C use. Further large-scale trials are warranted to elucidate its optimal intravenous dosage and effectiveness against chronic pain in the postoperative pain control setting.


High levels of human immunodeficiency virus infection of CD8 lymphocytes expressing CD4 in vivo.
Human immunodeficiency virus (HIV)-infected CD8 lymphocytes have been reported in vivo, but the mechanism of infection remains unclear. Experiments using the thy/hu mouse model support export of intrathymically infected CD8 precursors, while recent in vitro data suggest that mature CD8 lymphocytes upregulate CD4 upon activation (generating a CD8bright CD4dim phenotype) and are susceptible to HIV infection. To determine whether these mechanisms operate in vivo and to assess their relative importance in the generation of circulating HIV-infected CD8 lymphocytes, we quantified HIV long terminal repeat (LTR) DNA in CD8+ CD4- and CD8bright CD4dim lymphocytes isolated from HIV-infected individuals by fluorescence-activated cell sorting. HIV infection of CD8 lymphocytes was demonstrated in 17 of 19 subjects, with a significant inverse relationship between level of infection and CD4 lymphocyte count (R = -0.73; P < 0.001). The level of HIV infection of CD8bright CD4dim lymphocytes was significantly higher (median, 1,730 HIV LTR copies/10(6) cells; n = 9) than that of CD8+ CD4- lymphocytes (undetectable in seven of nine individuals; P < 0.01) and approached that of CD4 lymphocytes from the same individuals (median, 3,660 HIV LTR copies/10(6) cells). CD8bright CD4dim lymphocytes represented 0.8 to 3.3% of total CD8 lymphocytes and were most prevalent in the memory subset. Thus, HIV-infected CD8 lymphocytes commonly circulate in HIV-infected individuals and are generated through infection of activated CD8 lymphocytes rather than through export of intrathymically infected precursors. The high level of infection of CD8bright CD4dim lymphocytes could have a direct role in the decline in CD8 lymphocyte function that accompanies HIV disease progression.


Glucose homeostasis in rainbow trout fed a high-carbohydrate diet: metformin and insulin interact in a tissue-dependent manner.
Carnivorous fish species such as the rainbow trout (Oncorhynchus mykiss) are considered to be "glucose intolerant" because of the prolonged hyperglycemia experienced after intake of a carbohydrate-enriched meal. In the present study, we use this species to study glucose homeostasis in fish chronically infused with the hypoglycemic agents, insulin, and metformin, and fed with a high proportion of carbohydrates (30%). We analyzed liver, skeletal muscle, and white adipose tissue (WAT), which are insulin- and metformin-specific targets at both the biochemical and molecular levels. Trout infused with the combination of insulin and metformin can effectively utilize dietary glucose at the liver, resulting in lowered glycemia, increased insulin sensitivity, and glucose storage capacity, combined with reduced glucose output. However, in both WAT and skeletal muscle, we observed decreased insulin sensitivity with the combined insulin + metformin treatment, resulting in the absence of changes at the metabolic level in the skeletal muscle and an increased potential for glucose uptake and storage in the WAT. Thus, the poor utilization by rainbow trout of a diet with a high proportion of carbohydrate can at least be partially improved by a combined treatment with insulin and metformin, and the glucose intolerance observed in this species could be, in part, due to some of the downstream components of the insulin and metformin signaling pathways. However, the predominant effects of metformin treatment on the action of insulin in these three tissues thought to be involved in glucose homeostasis remain exclusive in this species.


Extracorporeal shockwave lithotripsy for lower pole calculi smaller than one centimeter.
Extracorporeal shockwave lithotripsy (ESWL) has revolutionized the treatment of urinary calculi and became the accepted standard therapy for the majority of stone patients. Only for stones located in the lower calix, ESWL displayed a limited efficacy. Since the stone-free rate seemed to be preferential, endoscopic maneuvers like percutaneous nephrolithotomy (PCNL) and retrograde intrarenal surgery (RIRS) have been proposed as the primary approach for this stone localization.Stone size seems to be the most important parameter in regard to the stone-free rate, whereas anatomical characteristics of the lower pole collecting system are discussed controversial. Various studies show a good stone clearance between 70-84% for stones up to 1 cm in diameter. Additional physical and medical measures are suitable to improve treatment results. Stone remnants after ESWL, defined as clinical insignificant residual fragments (CIRF) will not cause problems in every case and will pass until up to 24 months after treatment; in total 80-90% of all patients will become stone-free or at least symptom-free.When complete stone-free status is the primary goal, follow-up examinations with new radiological technologies like spiral CT show that the stone-free rate of ESWL and endoscopically treated patients (RIRS) does not differ significantly. However, in comparison to endoscopic stone removal, shockwave therapy is noninvasive, anesthesia-free and can be performed in an outpatient setup. Therefore, ESWL remains the first choice option for the treatment of lower caliceal stones up to 1 cm. The patient will definitely favour this procedure.


Fractionated, single-port radiotherapy delays paresis in a metastatic spinal tumor model in rats.
Spinal column metastatic disease affects thousands of cancer patients every year. Radiation therapy frequently represents the primary treatment for this condition. Despite the enormous clinical impact of spinal column metastatic disease, the literature currently lacks an accurate animal model for testing the efficacy of irradiation on spinal column metastases. After anesthesia was induced, female Fischer 344 rats underwent a transabdominal approach to the ventral vertebral body (VB) of L-6. A 2- to 3-mm-diameter bur hole was drilled for the implantation of a section of CRL-1666 breast adenocarcinoma. After the animals had recovered from the surgery, they underwent fractionated, single-port radiotherapy beginning on postoperative Day 7. Each group of animals underwent five daily fractions of radiation treatment. Group I animals received a total dose of 10 Gy in 200-cGy daily fractions, Group II animals received a total dose of 20 Gy in 400-cGy daily fractions, and Group III animals received a total dose of 30 Gy in 600-cGy daily fractions. A control group of rats with implanted VB lesions did not receive radiation. To test the effects of radiation toxicity alone, additional rats without implanted tumors received radiation treatments in the same fractions as the rats with tumors. Hindlimb function in all rats was rated before and after radiation treatment using the Basso-Beattie-Bresnahan locomotor rating scale. Histological analysis of spinal cord and vertebral column sections was performed after each animal's death. Functional assessments demonstrated a statistically significant delay in the onset of paresis between the three treatment groups and the control group (tumor implanted but no radiotherapy). The rats in the three treatment groups, however, did not exhibit any significant differences related to hindlimb function. A dose-dependent relationship was found for the percentage of animals who had become paralyzed at the time of death, with all members of the control group and no members of the 30-Gy group exhibiting paralysis. The results of this study do not indicate any overall survival benefit for any level of radiation dose. These findings demonstrate the efficacy of focal spinal irradiation in delaying the onset of paralysis in a rat metastatic spine tumor model, but without a clear survival benefit. Because of the dose-related toxicity observed in the rats treated with 30 Gy, this effect was most profound for the 20-Gy group. This finding parallels the observed clinical course of spinal column metastatic disease in humans and provides a basis for the future comparison of novel local and systemic treatments to augment the observed effects of focal irradiation.


Actin is required for IFT regulation in Chlamydomonas reinhardtii.
Assembly of cilia and flagella requires intraflagellar transport (IFT), a highly regulated kinesin-based transport system that moves cargo from the basal body to the tip of flagella [1]. The recruitment of IFT components to basal bodies is a function of flagellar length, with increased recruitment in rapidly growing short flagella [2]. The molecular pathways regulating IFT are largely a mystery. Because actin network disruption leads to changes in ciliary length and number, actin has been proposed to have a role in ciliary assembly. However, the mechanisms involved are unknown. In Chlamydomonas reinhardtii, conventional actin is found in both the cell body and the inner dynein arm complexes within flagella [3, 4]. Previous work showed that treating Chlamydomonas cells with the actin-depolymerizing compound cytochalasin D resulted in reversible flagellar shortening [5], but how actin is related to flagellar length or assembly remains unknown. Here we utilize small-molecule inhibitors and genetic mutants to analyze the role of actin dynamics in flagellar assembly in Chlamydomonas reinhardtii. We demonstrate that actin plays a role in IFT recruitment to basal bodies during flagellar elongation and that when actin is perturbed, the normal dependence of IFT recruitment on flagellar length is lost. We also find that actin is required for sufficient entry of IFT material into flagella during assembly. These same effects are recapitulated with a myosin inhibitor, suggesting that actin may act via myosin in a pathway by which flagellar assembly is regulated by flagellar length.


Persistent short nighttime sleep duration is associated with a greater post-COVID risk in fully mRNA-vaccinated individuals.
Short nighttime sleep duration impairs the immune response to virus vaccination, and long nighttime sleep duration is associated with poor health status. Thus, we hypothesized that short (<6 h) and long (>9 h) nighttime sleepers have a higher post-COVID risk than normal nighttime sleepers, despite two doses of mRNA vaccine (which has previously been linked to lower odds of long-lasting COVID-19 symptoms). Post-COVID was defined as experiencing at least one core COVID-19 symptom for at least three months (e.g., shortness of breath). Multivariate logistic regression adjusting for age, sex, BMI, and other factors showed in 9717 respondents (age span 18-99) that two mRNA vaccinations lowered the risk of suffering from post-COVID by about 21% (p < 0.001). When restricting the analysis to double-vaccinated respondents (n = 5918), short and long sleepers exhibited a greater post-COVID risk than normal sleepers (adjusted OR [95%-CI], 1.56 [1.29, 1.88] and 1.87 [1.32, 2.66], respectively). Among respondents with persistent sleep duration patterns during the pandemic compared to before the pandemic, short but not long sleep duration was significantly associated with the post-COVID risk (adjusted OR [95%-CI], 1.59 [1.24, 2.03] and 1.18 [0.70, 1.97], respectively). No significant association between sleep duration and post-COVID symptoms was observed in those reporting positive SARS-CoV-2 test results (n = 538). Our findings suggest that two mRNA vaccinations against SARS-CoV-2 are associated with a lower post-COVID risk. However, this protection may be less pronounced among those sleeping less than 6 h per night. Our findings warrant replication in cohorts with individuals with confirmed SARS-CoV-2 infection.


CD4+ T-cell immunity to mutated ras protein in pancreatic and colon cancer patients.
Mutated p21 ras proteins contain single substituted amino acid residues and represent cancer-specific proteins. The current study examined whether primed T cell immunity to mutant p21 ras proteins and/or peptides can be detected in patients with pancreatic or colon cancer. Studies focused on the aspartic acid substitution in amino acid position 12 (denoted D12) as the commonest mutation in gastrointestinal malignancy. Peripheral blood lymphocytes from patients or normal individuals were tested for the ability to proliferate in response to normal or mutated ras peptides or proteins. T-cell responses were defined as a stimulation index of > 2.0. Results showed that 7 of 16 (44%) pancreatic cancer patients responded to ras-D12 peptide. Responses to ras-D12 protein were studied in only the last four patients that responded to D12 peptides. Three of the 4 patients that responded to ras-D12 peptide showed a substantial response to p21 ras-D12 protein (stimulation indices of 12, 8, and 24). Specificity was validated by examining responses to normal and alternate ras peptides and proteins. T-cell responses to ras-D12 peptides were detected in only 2 of 25 (8%) colon cancer patients. None of 11 normal individuals tested had positive responses to normal or mutant ras p21 proteins and/or peptides. Thus, CD4+ T-cell immunity to the mutated segment of ras protein is present in some patients with gastrointestinal cancer.


Non-attendance rate in a Danish University Clinic of Dermatology.
To clarify the rate of non-attendance (NA) in an out-patient clinic. Attendance lists of 3592 patients were collected daily from 21 July-21 August and 21 October-21 November, 2009. NA patients were contacted to determine extenuations. To study NA in relation to diagnosis and age, a control group of patients who attended before or after a NA was established. Furthermore, two time periods from 8:00-11:30 AM and 11:30 AM-3:00 pm were compared. In total, 13% NA gave no cancellation (54.2% females and 45.8% males). Divided into age groups, 496 patients 0-25 years old had appointments, but 87 (18.6%) showed NA. In the 26-65 years old, 2188 patients were planned, but 313 (14.1%) showed NA. Over 65 years old, 878 patients were planned, but 69 patients (7.9%) showed NA. NA was higher (P < 0.05) in patients 0-25 years old in comparison with the other age groups. Diagnoses had no influence on the rate of NA (P > 0.05), neither had seasons nor time of the day. The main explanations reported by the NA were: forgetfulness (34.3%), erroneous scheduling (27.7%) and various reasons (38.0%). However, 18.5% had shown NA before while 17.1% were NA first timers. The NA rate 13% of 3592 patients was mostly patient-related. Erroneous scheduling was estimated to be 3.6%. NA was more frequent among young patients. NA rate is small in comparison with non-adherence to medicines, however, of major practical and economic consequence for the health system. SMS or e-mail notification and improved scheduling are potential actions to improve NA in the routine.


Activity-dependent PSA expression regulates inhibitory maturation and onset of critical period plasticity.
Functional maturation of GABAergic innervation in the developing visual cortex is regulated by neural activity and sensory inputs and in turn influences the critical period of ocular dominance plasticity. Here we show that polysialic acid (PSA), presented by the neural cell adhesion molecule, has a role in the maturation of GABAergic innervation and ocular dominance plasticity. Concentrations of PSA significantly decline shortly after eye opening in the adolescent mouse visual cortex; this decline is hindered by visual deprivation. The developmental and activity-dependent regulation of PSA expression is inversely correlated with the maturation of GABAergic innervation. Premature removal of PSA in visual cortex results in precocious maturation of perisomatic innervation by basket interneurons, enhanced inhibitory synaptic transmission, and earlier onset of ocular dominance plasticity. The developmental and activity-dependent decline of PSA expression therefore regulates the timing of the maturation of GABAergic inhibition and the onset of ocular dominance plasticity.


Unique mutations in mitochondrial DNA of senescence-accelerated mouse (SAM) strains.
Mitochondrial DNA (mtDNA) is exclusively inherited maternally and hence could offer a good method for tracing the lineage of mouse strains. We examined the mtDNA sequence of senescence-accelerated mouse (SAM) strains as well as other laboratory strains of inbred mice to deduce the ancestral strain of SAM. Four unique mutations were identified at bases 2256, 10,847, 11,181, and 13,053 in SAM strains. The mutations were not found in other mouse strains including AKR/J, one of the parental strains of SAM. Comparison of the mtDNA sequences also led to the consensus mtDNA sequence of laboratory strains of inbred mice. The seven laboratory strains of common inbred mice showed polymorphisms at base 9348, thymine repeat from base 9818, and adenine repeat from base 9821, and could be classified into five types by combination of the differences. Although we could not identify mouse strains with the same type of mtDNA as SAM in this study, the polymorphisms would provide a promising clue to ascertain the ancestral strain(s) of SAM. The polymorphism in mtDNA could be used to ascertain the genealogy of other mouse strains as well.


Pathogenesis of myonecrosis induced by coral snake (Micrurus nigrocinctus) venom in mice.
The mode by which coral snake (Micrurus nigrocinctus) venom affects skeletal muscle was studied using a combined approach. The venom induced early functional and structural alterations in the plasma membrane of muscle cells, suggesting that sarcolemma is the primary site of action of this venom. This was shown by the presence of wedge-shaped ('delta') lesions at the periphery of the cells, as well as by focal disruptions in the continuity of plasma membrane as early as 15 min after envenomation. After this initial alteration the rest of the organelles were severely affected. Myofilaments were hypercontracted leaving, as a consequence, areas of overstretched myofibrils as well as empty spaces. Eventually, myofilaments formed dense, clumped masses in which the striated structure was totally lost. At 24 h, myofilaments were still disorganized but they presented a more hyaline and homogeneous appearance. As early as 15 and 30 min mitochondria were swollen; later, by I, 3 and 24 h, they showed further alterations such as the presence of dense intracristal spaces and vesiculated cristae, as well as disruption in the integrity of their membranes. Sarcoplasmic reticulum was dilated and disorganized into many small vesicles randomly distributed throughout the cellular space. Moreover, the venom induced a rapid decrease in muscle levels of creatine and creatine-kinase (CK) and a calcium influx. Since the rates of efflux of creatine and CK were similar, it is suggested that the lesions produced in the membrane are large enough to allow the escape of these two molecules. As corroboration of the severe myotoxic effect, envenomated mice excreted reddish urine containing large quantities of myoglobin. Skeletal muscle cells are more susceptible to the action of the venom than erythrocytes, since coral snake venom induced only a mild direct haemolytic effect in vitro and haemolysis is not a significant effect in vivo. M. nigrocinctus venom induced a drastic increase in plasma levels of lactate dehydrogenase. Isozymes LDH-3, LDH-4, and LDH-5 increased markedly, suggesting that the systemic pathology of coral snake envenoming may be more complex than previously thought.


Chronic nicotine in hearts with healed ventricular myocardial infarction promotes atrial flutter that resembles typical human atrial flutter.
The potential of chronic nicotine exposure for atrial fibrillation (AF) and atrial flutter (AFL) in hearts with and without chronic myocardial infarction (MI) remains poorly explored. MI was created in dogs by permanent occlusion of the left anterior descending coronary artery, and dogs were administered nicotine (5 mg.kg(-1).day(-1) sc) for 1 mo using osmotic minipumps. High-resolution epicardial (1,792 bipolar electrodes) and endocardial Halo catheters were used to map activation during induced atrial rhythms. Nicotine promoted inducible sustained AFL at a mean cycle length of 134 +/- 10 ms in all MI dogs (n = 6) requiring pacing and electrical shocks for termination. No AFL could be induced in MI dogs (n = 6), control (non-MI) dogs (n = 3) not exposed to nicotine, and dogs with no MI and exposed to nicotine (n = 3). Activation maps during AFL showed a single reentrant wavefront in the right atrium that rotated either clockwise (60%) or counterclockwise (40%) around the crista terminalis and through the isthmus. Ablation of the isthmus prevented the induction of AFL. Nicotine caused a significant (P < 0.01) but highly heterogeneous increase in atrial interstitial fibrosis (2- to 10-fold increase in left and right atria, respectively) in the MI group but only a 2-fold increase in the right atrium in the non-MI group. Nicotine also flattened (P < 0.05) the slope of the epicardial monophasic action potential duration (electrical restitution) curve of both atria in the MI but not in non-MI dogs. Two-dimensional simulation in an excitable matrix containing an isthmus and nicotine's restitutional and reduced gap junctional coupling (fibrosis) parameters replicated the experiments. Chronic nicotine in hearts with MI promotes AFL that closely resembles typical human AFL. Increased atrial interstitial fibrosis and flattened electrical restitution are important substrates for the AFL.


Paediatric nephrology at the University Hospital of the West Indies. A walk through time.
Paediatric nephrology at the University Hospital of the West Indies has grown over the last five decades into an established paediatric subspecialty offering to Jamaica and other Caribbean territories the benefit of paediatricians with training and exposure in this field. Dissemination of information to medical practitioners within the island has reduced mortality and morbidity associated with potentially treatable atypical renal disease. Clear investigative guidelines for urinary tract infection have made earlier diagnosis of urological malformations possible. Patterns of glomerular disease in Jamaican children have now been aetiologically and histologically documented. Children with chronic kidney disease now have clear management guidelines and the possibility of renal replacement therapy. Future goals include community education about renal disease and the development of a paediatric dialysis and transplantation unit.